MOLECULAR ANALYSIS OF THE BOSS TRANSMEMBRANE LIGAND

细胞间的通讯在细胞分化中起着关键作用 流程。果蝇R7光感受器细胞的诱导 由邻近的R8细胞组成的复眼已成为经典之一 两个细胞之间的感应相互作用的例子。R7被诱导 通过无七(SEV)受体酪氨酸激酶之间的相互作用 在R7前体细胞和Seven less新娘(BOSS)的表面， 在邻近的R8细胞表面表达。长期目标 这项研究的目的是了解R7中的生化步骤 归纳途径。 这一提议集中在对诱导配体BOSS的分析上。 BOSS蛋白由一个很大的胞外区(498个氨基酸)组成 酸)，具有七个膜跨段(7TM)的跨膜区 Seven less的激活和R7的诱导是必需的区域 细胞的命运。大的胞外进入邻近的R7细胞作为 它与SIV受体相互作用的结果。这些发现 提出一些重要的问题：BOSS如何充当 SIV受体？7TM结构域是否构成结合的一部分 SEV受体的位置？或者，7TM区域是否有必要 与第二种蛋白质相互作用形成同源二聚体或异源二聚体？什么 BOSS转移到R7的机制是什么？ 这些问题将通过以下具体目标加以解决： 1.确定BOSS 7TM区域在激活SEV受体中的作用 酪氨酸激酶。嵌合蛋白将被用来定义最小的 R7诱导所需的7TM区域的一部分，并推断其 功能意义。 2.对BOSS蛋白进行纯化和功能重组。如果 重组BOSS蛋白不足以激活SIV 受体酪氨酸激酶我们将鉴定BOSS的中间步骤 内部化。 信号的许多--如果不是全部--关键成分的突变 诱导R7发育的转导途径很快就会出现。 这些突变为分析这一现象提供了完美的环境 路径。使用细胞的生物化学和遗传学方法的组合 培养系统和转基因动物将允许亲密的知识 在这条道路上。这一知识将为 了解调控如此多样化的多种信号通路 神经分化、生长控制和肿瘤形成等过程。一个 在特定蛋白质-蛋白质水平上的信号转导知识 互动将是朝着 合理设计调节这些过程的药物。

Cell-cell communication plays a key role in may cellular differentiation processes. Induction of the R7 photoreceptor cell in the Drosophila compound eye by the neighboring R8 cell has become one of the classic examples for an inductive interaction between two cells. R7 is induced by an interaction between the sevenless (sev) receptor tyrosine kinase on the surface of the R7 precursor cell and bride of sevenless (boss), expressed on the surface of the neighboring R8 cell. The long term goal of this research is to understand the biochemical steps in the R7 inductive pathway. This proposal concentrates on an analysis of the inducing ligand boss. The boss protein is composed of a large extracellular domain (498 amino acids), a transmembrane region with seven membrane spanning segments (7TM region is necessary for the activation of sevenless and induction of R7 cell fate. The large extracellular into the neighboring R7 cell as a consequence of its interaction with the sev receptor. These findings raise a number of important questions: How does boss act as a ligand for the sev receptor? Does the 7TM domain constitute part of the binding site for the sev receptor? Alternatively, is the 7TM region necessary to interact with a second protein to form a homodimer or heterodimer? What is the mechanism of transfer of boss into R7? These questions will be addressed by the following specific aims: 1. Define the role of the boss 7TM region in activating the sev receptor tyrosine kinase. Chimeric proteins will be used to define the minimal part of the 7TM region required for R7 induction and to infer its functional significance. 2. Purify and functionally reconstitute the boss protein. If reconstituted boss protein is not sufficient for activation of the sev receptor tyrosine kinase we will identify intermediate steps of boss internalization. Mutations in many -if not all- of the critical components of the signal transduction pathway inducing R7 development will soon be available. These mutations provide a perfect setting for an analysis of this pathway. A combination of biochemical and genetic methods using cell culture systems and transgenic animals will allow an intimate knowledge of this pathway. This knowledge will lay the foundation for understanding numerous signaling pathways regulating such diverse processes as neuronal differentiation, growth control and neoplasia. A knowledge of signal transduction at the level of specific protein-protein interactions will be an invaluable step forward in the direction of the rational design of drugs regulating these processes.